Identification of molecular-mimicry-based ligands for cholera diagnostics using magnetic relaxation

When covalently bound to an appropriate ligand, iron oxide nanoparticles can bind to a specific target of interest. This interaction can be detected through changes in the solution's spin-spin relaxation times (T2) via magnetic relaxation measurements. In this report, a strategy of molecular mimicry was used in order to identify targeting ligands that bind to the cholera toxin B subunit (CTB). The cellular CTB-receptor, ganglioside GM1, contains a pentasaccharide moiety consisting in part of galactose and glucose units. We therefore predicted that CTB would recognize carbohydrate-conjugated iron oxide nanoparticles as GM1 mimics, thus producing a detectable change in the T2 relaxation times. Magnetic relaxation experiments demonstrated that CTB interacted with the galactose-conjugated nanoparticles. This interaction was confirmed via surface plasmon resonance studies using either the free or nanoparticle-conjugated galactose molecule. The galactose-conjugated nanoparticles were then used as CTB sensors achieving a detection limit of 40 pM. Via magnetic relaxation studies, we found that CTB also interacted with dextran-coated nanoparticles, and surface plasmon resonance studies also confirmed this interaction. Additional experiments demonstrated that the dextran-coated nanoparticle can also be used as CTB sensors and that dextran can prevent the internalization of CTB into GM1-expressing cells. Our work indicates that magnetic nanoparticle conjugates and magnetic relaxation detection can be used as a simple and fast method to identify targeting ligands via molecular mimicry. Furthermore, our results show that the dextran-coated nanoparticles represent a low-cost approach for CTB detection.     

Biochemical Evaluation of Triploid Progenies of Diploid × Tetraploid Breeding Populations of Camellia for Genotypes Rich in Catechin and Caffeine

To verify the quality of triploid varieties of Camellia tea species at the secondary metabolite level, we tested caffeine and catechin profiles of 97 F₁ segregating progenies in two breeding populations with a common tetraploid parent and diploid parents of two geographic and varietal origins. Catechin and caffeine levels of the triploid progenies were quantified and compared against their diploid parent. Some of the progenies showed better performance than their diploid parent. Most of the progenies of the diploid C. sinensis × tetraploid cross showed heterosis for caffeine and EGCG. Progenies of the C. assamica subsp. lasiocalyx × tetraploid cross showed heterosis for +C, EC, EGC, and TC. The genomic contributions of the diploid parent seem to be the main factor in the variation between the two populations. Our studies showed quantitative enhancement of some of the quality-related parameters in tea, providing a platform to refocus on this classical breeding approach for developing quality cultivars in tea.     

Development of chitosan-tripolyphosphate fibers through pH dependent ionotropic gelation

Incorporation of phosphate groups into a material may be of particular interest as they act as templates for hydroxyapatite growth through complexation with Ca²⁺ and thus improve the osteoconduction property. The phosphate groups can be incorporated into chitosan through ionotropic gelation with tripolyphosphate (TPP). Interestingly, the ion pairs formed through negatively charged phosphate groups with protonated amine functionality of chitosan in ionotropic gelation are expected to provide chitosan with an amphoteric character, which may facilitate protein adhesion following enhanced attachment of anchorage dependant cells than chitosan, which shows poor cell adhesion properties. In this study, chitosan–tripolyphosphate (TPP) fibers with varying phosphate contents were prepared through wet spinning in STPP baths of different pH. Gelation kinetics and gel strength of chitosan with STPP solutions of three different pH were evaluated and compared with that of NaOH solution for evaluation of their influence on nature of gelation. The solution pH of STPP baths was found to have significant control on the extent of ionic cross-linking and physico-chemical properties of the fibers. Moreover, this kinetically driven ionotropic gelation of chitosan by TPP results in low degree of crystallinity of chitosan–TPP fibers and consequently their lower thermal stability than chitosan fibers.     

Exploring the Zika Genome to Design a Potential Multiepitope Vaccine Using an Immunoinformatics Approach

International Journal of Peptide Research and Therapeutics - Zika is one of the most dreaded viruses which has left mankind crippled for over years. Current no vaccines for Zika are available in...     

A Novel Role for Lh3 Dependent ECM Modifications during Neural Crest Cell Migration in Zebrafish

During vertebrate development, trunk neural crest cells delaminate along the entire length of the dorsal neural tube and initially migrate as a non-segmented sheet. As they enter the somites, neural crest cells rearrange into spatially restricted segmental streams. Extracellular matrix components are likely to play critical roles in this transition from a sheet-like to a stream-like mode of migration, yet the extracellular matrix components and their modifying enzymes critical for this transition are largely unknown. Here, we identified the glycosyltransferase Lh3, known to modify extracellular matrix components, and its presumptive substrate Collagen18A1, to provide extrinsic signals critical for neural crest cells to transition from a sheet-like migration behavior to migrating as a segmental stream. Using live cell imaging we show that in lh3 null mutants, neural crest cells fail to transition from a sheet to a stream, and that they consequently enter the somites as multiple streams, or stall shortly after entering the somites. Moreover, we demonstrate that transgenic expression of lh3 in a small subset of somitic cells adjacent to where neural crest cells switch from sheet to stream migration restores segmental neural crest cell migration. Finally, we show that knockdown of the presumptive Lh3 substrate Collagen18A1 recapitulates the neural crest cell migration defects observed in lh3 mutants, consistent with the notion that Lh3 exerts its effect on neural crest cell migration by regulating post-translational modifications of Collagen18A1. Together these data suggest that Lh3–Collagen18A1 dependent ECM modifications regulate the transition of trunk neural crest cells from a non-segmental sheet like migration mode to a segmental stream migration mode.     

Dry-reagent gold nanoparticle-based lateral flow biosensor for the simultaneous detection of Vibrio cholerae serogroups O1 and O139

Cholera is a communicable disease caused by consumption of contaminated food and water. This potentially fatal intestinal infection is characterised by profuse secretion of rice watery stool that can rapidly lead to severe dehydration and shock, thus requiring treatment to be given immediately. Epidemic and pandemic cholera are exclusively associated with Vibrio cholerae serogroups O1 and O139. In light of the need for rapid diagnosis of cholera and to prevent spread of outbreaks, we have developed and evaluated a direct one-step lateral flow biosensor for the simultaneous detection of both V. cholerae O1 and O139 serogroups using alkaline peptone water culture. Serogroup specific monoclonal antibodies raised against lipopolysaccharides (LPS) were used to functionalize the colloidal gold nanoparticles for dual detection in the biosensor. The assay is based on immunochromatographic principle where antigen-antibody reaction would result in the accumulation of gold nanoparticles and thus, the appearance of a red line on the strip. The dry-reagent dipstick format of the biosensor ensure user-friendly application, rapid result that can be read with the naked eyes and cold-chain free storage that is well-suited to be performed at resource-limited settings.     

Cse4 (CenH3) association with the Saccharomyces cerevisiae plasmid partitioning locus in its native and chromosomally integrated states: implications in centromere evolution

The histone H3 variant Cse4 specifies centromere identity in Saccharomyces cerevisiae by its incorporation into a special nucleosome positioned at CEN DNA and promotes the assembly of the kinetochore complex, which is required for faithful chromosome segregation. Our previous work showed that Cse4 is also associated with the partitioning locus STB of the 2μm circle--a multicopy plasmid that resides in the yeast nucleus and propagates itself stably. Cse4 is essential for the functional assembly of the plasmid partitioning complex, including the recruitment of the yeast cohesin complex at STB. We have located Cse4 association strictly at the origin-proximal subregion of STB. Three of the five directly repeated tandem copies of a 62-bp consensus sequence element constituting this region are necessary and sufficient for the recruitment of Cse4. The association of Cse4 with STB is dependent on Scm3, the loading factor responsible for the incorporation of Cse4 into the CEN nucleosome. A chromosomally integrated copy of STB confers on the integration site the capacity for Cse4 association as well as cohesin assembly. The localization of Cse4 in chromatin digested by micrococcal nuclease is consistent with the potential assembly of one Cse4-containing nucleosome, but not more than two, at STB. The remarkable ability of STB to acquire a very specialized, and strictly regulated, chromosome segregation factor suggests its plausible evolutionary kinship with CEN.     

Spectroscopic, cytotoxic and DFT studies of a luminescent palladium(II) complex of a hydrazone ligand that induces apoptosis in human prostate cancer cells

A luminescent palladium(II) complex [Pd(L)Cl], 1a was synthesized with the acyclic tridentate quinoline-2-carboxaldehyde-2-pyridylhydrazone ligand, HL, 1. The ligand, 1 showed a selective chromogenic behavior towards Pd²⁺ by changing the color of the solution from yellow to blue-violet, which can easily be detected by the naked-eye. DFT and TDDFT calculations were performed to determine the geometry optimized structures of the ligand 1 and the complex 1a as well as to correlate the electronic transitions. The complex 1a exhibits strong interaction towards DNA as revealed from K_b (intrinsic binding constant) and K_sq (Stern Volmer quenching constant) values, which are 1.47 × 10⁵ M⁻¹ and 5.67, respectively. The cytotoxicity of 1a has been examined with human prostate cancer cells (PC-3) and the sub-lethal dose (8 μM) determined by dose-dependence studies. The relative degree of apoptotic and necrotic cell death using a sub-lethal dose were measured by flow cytometry. The cell cycle analysis shows that the complex 1a exhibits effective cell growth inhibition by triggering G2/M phase arrest and apoptosis in cancer cells. Moreover, its treatment triggers the mitochondrial pathway resulting in cytochrome c release and caspase-3 activation.